骨髓间充质干细胞向多巴胺能神经元的诱导分化  

Induction and differentiation of bone marrow mesenchymal stem cells into dopaminergic neurons

作　　者：刘卓[1] 徐运[1] 黄丹青[1] 

机构地区：[1]南京大学医学院附属鼓楼医院神经内科,江苏省南京市210008

出　　处：《中国组织工程研究与临床康复》2010年第23期4227-4230,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：南京大学医学院附属鼓楼医院青年科研启动基金;课题名称:骨髓间充质干细胞诱导为神经细胞的实验研究~~

摘　　要：背景:近年来研究发现,神经营养因子在骨髓间充质干细胞的分化中发挥重要作用。目前脑组织中具有再生能力的神经干细胞在体外是否具有直接诱导骨髓间充质干细胞分化为多巴胺能神经元的作用还未见报道。目的:观察大鼠间充质干细胞在胶质细胞源性神经营养因子与神经干细胞共培养两种诱导条件下体外分化成多巴胺能神经元的能力。方法:分离培养SD大鼠骨髓间充质干细胞,取第3代细胞分2组培养,一组细胞应用胶质细胞源性神经营养因子单独诱导,另一组细胞与已培养成球的神经干细胞共培养进行诱导,共培养之前行Brdu标记。诱导3d后以免疫组织化学法检测各组贴壁细胞神经元特异性标志物神经原纤维和多巴胺能神经元特异性标志物酪氨酸羟化酶的表达,观察间充质干细胞的分化情况。结果与结论:胶质细胞源性神经营养因子单独诱导组间充质干细胞在诱导24h后胞体回缩呈锥形,突起延长且数量增多,有神经元样形态,且细胞间相互连接成网络状,3d后部分细胞表达神经原纤维,其中少部分同时表达酪氨酸羟化酶。与神经干细胞共培养组神经干细胞球很快解离,迅速贴壁,共培养的贴壁细胞大量增殖且多呈神经元样,胞体细长多突起,相互间连接成网,多数贴壁细胞分别单独表达神经原纤维和酪氨酸羟化酶,少数细胞可见Brdu/神经原纤维,Brdu/胶质纤维酸性蛋白,Brdu/酪氨酸羟化酶双标阳性。提示间充质干细胞在胶质细胞源性神经营养因子、神经干细胞存在的情况下可定向转化为神经元,并有向多巴胺能神经元分化的可能。在该实验条件下胶质细胞源性神经营养因子效果好于神经干细胞。BACKGROUND：Recent studies have found that neurotrophic factor plays important roles in differentiation of bone mesenychymal stem cells （BMSCs）.However,few reports have addressed whether neural stem cells （NSCs） with regeneration capacity in the brain tissue in vitro can induce the differentiation of BMSCs into dopaminergic neurons.OBJECTIVE：To investigate the possibility of rat BMSCs differentiation into dopaminergic neurons under the induction condition of glial cells-derived neurotrophic factor （GDNF） and NSCs coculture.METHODS：BMSCs from Sprague Dawley rats were isolated and cultured.The third passage of BMSCs was divided into two groups.Group A：Medium was MEM containing GDNF;group B：BMSCs marked with Brdu and then cocultured with NSCs which had formed primary neurospheres.Three days later,BMSCs from each group were suffered immunohistochemistry staining to examine the tyrosine hydroxylase of the differentiated neuron to observe the differentiation of BMSCs.RESULTS AND CONCLUSION：At 24 hours postinduction,BMSCs of group A were cone-shaped and had the prolonged processes.The amount of processes increased,with the presence of neuron-like morphology.The web connection among the cells can also be observed.Three days later,some cells expressed tyrosine hydroxylase.In group B,NSCs attached to culture dish immediately.The attached cells proliferated and showed neuronal phenotype.A majority of attached cells expressed tyrosine hydroxylase,and a few cells presented double staining：Brdu/tyrosine hydroxylase.Results have exhibited that BMSCs have the potential of differentiating into dopaminergic neurons.The GDNF proves to be more effective in the experiment compared with NSCs.

关 键 词：骨髓间充质干细胞 神经干细胞 胶质细胞源性神经营养因子 诱导 神经元 

分 类 号：R394.2[医药卫生—医学遗传学]